Reticulitermes flavipes termites synthesize uric acid via purine-nucleoside phosphorylase (purine-nucleoside: orthophosphate ribosyltransferase, EC 2.4.2.1) and xanthine dehydrogenase (xanthine:NAD' oxidoreductase, EC 1.2.1.37), but their tissues lack uricase (urate:oxygen oxidoreductase, EC 1.7.3.3) or any other enzyme that degrades uric acid. Nevertheless, uricolysis occurs in termites, but as an anaerobic process mediated by hindgut bacteria. "4C-Tracer experiments showed that termites transport uric acid from the site ofsynthesis and storage (fat body tissue) to the site ofdegradation (hindgut microbiota) via Malpighian tubules. Moreover, [1,3-'5N]uric acid dissimilated by gut bacteria in vivo leads to assimilation of 15N into termite tissues. NH3, a product of uricolysis, is a potential N source for termites, either directly via glutamine synthetase [L-glutamate: ammonia ligase (ADP-forming), EC 6.3.1.2] activity of fat body tissue or indirectly through microbe assimilation. Symbiotic recycling of uric acid N appears to be important to N conservation in these oligonitrotrophic insects.Wood-eating termites are oligonitrotrophic: their diets contain only 0.03-0.15% N and have a C/N ratio of 400:1 (1, 2). For such animals, conservation of N is crucial, and an intriguing strategy to accomplish this was suggested in 1938 by Leach and Granovsky (3). They speculated that the N atoms of uric acid, a common excretory product of terrestrial insects, might be recycled back to termites through the action of uricolytic gut microbes. For more than 40 years this provocative hypothesis remained untested. However, recent work in our laboratory revealed that (i) termites contain uric acid, but they do not void the purine despite the apparent absence of uricase (urate oxidase, urate:oxygen oxidoreductase, EC 1.7.3.3) in their tissues (4) and (ii) the hindgut of Reticulitermes flavipes harbors uricolytic bacteria capable of fermenting uric acid to C02, NH3, and acetate (5, 6). It remained to be shown, however, that termites transport uric acid to the hindgut microbiota and that termites derive nutritional benefit from microbial uricolysis. Clarification ofthese issues constitutes a major aspect ofthe present paper. MATERIALS AND METHODSInsects and Bacterial Strains. R. flavipes worker termites were used. They were collected in Michigan and Wisconsin and were maintained as described (7). Drosophila melanogaster was obtained from T. Friedman of Michigan State University.Streptococcus sp. UAD-1 and Bacteroides termitidis UAD-50 were among the predominant uricolytic bacteria isolated from guts of R. flavipes (5, 6).Preparation ofTissues, Cell Suspensions, and Cell Extracts. Termite Malpighian tubules were obtained by removing the intestinal tract (8) into 0.25 M sucrose/1.7 mM EDTA, pH 6.9, and dissecting the tubules free. Fat body tissue was scraped from the abdominal wall of degutted termites into 0.5 ml of sucrose/EDTA.Abdomen extracts were an easily obtainable, but crude, source of fat body tissue enzymes. Degutted a...

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Two strains of facultatively anaerobic, N,-fixing bacteria were isolated from guts of Coptotermes formosanus and identified as Enterobacter agglomerans. The deoxyribonucleic acid base composition of isolates was 52.6 and 53.1 mol% guanine plus cytosine. Both isolates and a known strain of E. agglomerans carried out a mixed acid type of glucose fermentation. N2 fixation by E. agglomerans was inhibited by 02; consequently, N. served as an N source only for cells growing anaerobically in media lacking a major source of combined N. However, peptone, NH4Cl, or KNO:, served as an N source under either aerobic or anaerobic conditions. It was estimated that 2 x 102 cells of E. agglomerans were present per termite gut. This value was 100-fold lower than expected, based on N. fixation rates of E. agglomerans in vitro and that of the intact termites. However, low recoveries of E. agglomerans may be related to the marked decrease in N, fixation rates observed when intact termites or their extracted guts were manipulated for the isolation of bacteria. It was concluded that the N2fixing activity of E. agglomerans may be important to the N economy of C. formosanus.

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A study was done of anaerobic degradation of uric acid (UA) by representative strains of uricolytic bacteria isolated from guts of Reticulitermes flavipes termites. Streptococcus strain UAD-1 degraded UA incompletely, secreting a fluorescent compound into the medium, unless formate (or a formicogenic compound) was present as a cosubstrate. Formate functioned as a reductant, and its oxidation to CO2 by formate dehydrogenase provided 2H+ + 2e-needed to drive uricolysis to completion. Uricolysis by Streptococcus UAD-1 thus corresponded to the following equation: 1UA + lformate-. 4CO2 + lacetate + 4NH3. Urea did not appear to be an intermediate in CO2 and NH3 formation during uricolysis by strain UAD-1. Formate dehydrogenase and uricolytic activities of strain UAD-1 were inducible by growth of cells on UA. Bacteroides ternitidis strain UAD-50 degraded UA as follows: 1UA-* 3.5 CO2 + 0.75acetate + 4NH3. Exogenous formate was neither required for nor stimulatory to uricolysis by strain UAD-50. Studies of UA catabolism by Citrobacter strains were limited, because only small amounts of UA were metabolized by cells in liquid medium. Uricolytic activity of such bacteria in situ could be important to the carbon, nitrogen, and energy economy of R. flavipes.